Bioactive Benzocycloheptene Analogues From Himachalenes and its Applications

ABSTRACT

Functionalized benzocycloheptenes are one of the most important classes of bicyclic framework that have been investi-gated in different areas of biological activities. The claimed invention provides preparations of benzocycloheptene analogues, inhibitors of PI3K and MK2, pharmaceutical compositions containing them and their use in therapy. The compounds of Formula I, II, III, IV, V and VI may be used as anti type 2 diabetes, antipyretic, anti-inflammatory, antiepileptic, anticancer, antiulcer, CNS-stimulant, and CNS-depressant. These benzocycloheptene derivatives are useful in treatment of PI3K and MK2 related disorders.

FIELD OF THE INVENTION

The present invention relates to new compounds of formula (I), (II),(III), (IV), (V), (VI) and (VII) wherein R¹, R², R³, R⁴, R⁵ and X, Y andZ are as defined herein, and their applications in treatment of PI3K andMK2 related disorders for use as antipyretic, anti-inflammatory,antiepileptic, anticancer, antiulcer diseases, CNS-stimulant, andCNS-depressant activities and in the manufacture of medicines/drugs forsuch treatment.

BACKGROUND OF THE INVENTION

Benzocycloheptene is a bicyclic skeleton that comprises of ringformation as one of the pivotal transformation [Y. Liu, J. Su, J. H.Xiao, S. B. Jiang, H. Lu, W. Zhong, L. L. Wang, X. H. and S. Li, ChineseChem. Lett., 2008, 19, 428-430; W. Adam, M. Balci, Z. Ceylan and R. F.,Chem. Ber., 1991, 124, 383-386]. Benzocycloheptene and their derivativesare attractive biological targets in theoretical chemistry andpharmaceutical sciences, and coordination chemistry [V. K. Tandon, K. A.Singh, A. K. Awasthi, J. M. Khanna, B. Lal and N. Anand, Bioorg. Med.Chem. Lett., 2004, 14, 2867-2870; M. Shiraishi, Y. Aramaki, M. Seto, H.Imoto, Y. Nishikawa, N. Kanzaki, M. Okamoto, H. Sawada, O. Nishimura, M.Baba and M. Fujino, J Med Chem., 2000 43, 2049-2063; R. Wyrwa, O.Peters, R. Bohlmann, P. Droescher, K. Prellle, K. H. Fritzemeier, H. P.Muhn, Patent US2009/0099250 A1, 2009]. Among benzocycloheptene, aminesubstituted benzocycloheptene have drawn much attention and intenseresearch activity because of their profound pharmacological potential.These were reported for the treatment of cancer [M. Sriram, J. J. Hall,N. C. Grohmann, T. E. Strecker, T. Wootton, A. Franken, M. L. Trawickand K. G. Pinney, Bioorg. Med. Chem., 2008, 16, 8161-8171; H. Strobel,P. Wohlfart, U.S. Pat. No. 6,759,412 B2, 2004; Z. Chen, C. J. O'Donnelland A. Maderna, Tetrahedron Lett., 2012, 53, 64-66], mental disorder [M.Nakano, M. Minoguchi, T. Hanano, S.-I. Ono, H. Horiuchi, K. Teshima, USPatent, 0120841A1, 2010], cardiovascular [CV. Amsterdam, PatentApplication no. WO/2002/030422A1, 2002], neurodegenerative [K. Kato, J.Terauchi, H. Fukumoto, M. Kakihana, Patent U.S. Pat. No. 7,256,204B2,2007], antidepressant [L. Nedelec, A. Pierdet, C. Dumont, M-H.Kannengiesser, Patent U.S. Pat. No. 4,148,919, 1979] and also work asantiarrhythmic [M. Baumgarth, I. Lues, K-O Minck,. N. Beier, Patent U.S.Pat. No. 5,495,022, 1996] and antiparkinson agents. The arylatedbenzosuberene derivatives is established in medicinal chemistry asanticancer agents because of their structural reminiscence to colchicineand combretastatin CA4 and CA1 [G. R. Pettit, S. B. Singh, C. M.Hamel-Lin, D. S. Alberts and D. Garcia-Kendall, Experientia., 1989, 45,209]. The major synthetic methods for benzocycloheptene synthesissuffered from some disadvantages like the tedious reaction conditions[M. Sriram, J. J. Hall, N. C. Grohmann, T. E. Strecker, T. Wootton, A.Franken, M. L. Trawick and K. G. Pinney, Bioorg. Med. Chem., 2008, 16,8161-8171.], difficulty to prepare the substituted congeners, thecommercial available starting materials [R. P. Tanpure, C. S. George, M.Sriram, T. E. Strecker, J. K. Tidmore, E. Hamel, A. K. Charlton-Sevcika,D. J. Chaplin, M. L. Trawick and K. G. Pinney, Med. Chem. Commun., 2012,6, 720], and the low yields [P. Y. Maximov, D. J. Fernandes, R. E.McDaniel, C. B. Myers, R. F. Curpan and V. C. Jordan, J. Med. Chem.,2014, 57, 4569]. In organic chemistry, chemistry of heterocycliccompounds is an interesting area of research since last decades. Variousheteroatoms like oxygen [Liu, R. S. PURE APPL CHEM., 2001, 73, 265-269],phosphorus [G. P. V. Reddy, Y. B. Kiran, S. C. Reddy and D. C. Reddy,Chem. Pharm. Bull., 2004, 52, 307-310], sulfur, nitrogen [M. G. Valverdeand T. Torroba, Molecules., 2005, 10, 318-320] and selenium [A. Hafez,Eur. J. Med. Chem., 2008, 43, 1971-1977] containing heterocycles arevery attention grabbing. However, among them, N-containing heterocycliccompounds have maintained the interest of researchers through decades ofhistorical development of organic synthesis. According to FDA database,the nitrogen-based heterocycles shows their structural significance inthe drug design and engineering of pharmaceuticals, with nearly 60% ofunique small-molecule drugs containing nitrogen heterocycles [E. Vitaku,D. T. Smith and J. T. Njardarson, J. Med. Chem., 2014, 57,10257-10274.]. Moreover, benzocycloheptene moieties contain fused sixand seven membered ring system, their derivatives possess potentialbacteriostatic, antipyretic, anti-inflammatory, anti-ulcer,CNS-stimulant, CNS-depressant and anti-convulsant activities [B. J.Crielaard, S. Van der Wal, T. Lammers, H. T. Le, W. E. Hennink, R. M.Schiffelers, G. Storm and M. H. A. M. Fens, Int. J. Nanomed., 2011, 6,2697-2703]. Some of the benzocyclohepta bicycle fused to nitrogenheterocycles derivatives present interesting pharmaceutical activities.Thus, 6,11-dihydro-5H-benzo[5,6]cyclohepta-[1,2-b]pyridine andN-substituted benzo[5,6]cyclohepta[1,2-b]indole are potent antitumoragents [A. Afonso, J. M. Kelly, J. Weinstein, R. L. Wolin, S. B.Rosenblum, U.S. Pat. No. 6,218,401. 2001, Chemical Abstract 134, 295746;J. Benoit, D. Alagille, J.-Y. Merour and S. Leonce, Chem. Pharm. Bull.,2000, 48, 1872-1876] against L1210 murine leukemia and HT29 cell lines,while piperazinyl and piperidinyl derivatives inhibit the farnesylprotein transferase (FPT) [S. W. Remiszewski, R. J. Doll, C. Alvarez, T.Lalwani, U.S. Pat. No. 6,211,193, 2001. Chemical Abstract 134, 266206].Synthesis of fused heterocyclic benzocycloheptene derivatives andefficiently execution these synthesized molecules into bioactivemolecules is an attention grabbing topic in medicinal chemistry.Sulfones are valuable synthetic targets [M. N. Noshi, A. El-awa, E.Torres and P. L. Fuchs, J. Am. Chem. Soc., 2007, 129, 11242-11247; J. N.Desrosiers and A. B. Charette, Angew Chem. Int. Ed., 2007, 46,5955-5957;. G. Pandey, K. N. Tiwari and V. G. Puranik, Org. Lett., 2008,10, 3611-3614; K. Oh. Org. Lett., 2007, 9, 2973-2975] due to versatilityof sulfone moieties [T. Nishimura, Y. Takiguchi and T. Hayashi, J. Am.Chem. Soc., 2012, 134, 9086-9089; J. N. Desrosiers and W. S. Bechara,Org. Lett., 2008, 10, 2315-2318]. It is a significant component innaturally occurring products and in drug discovery [U. Mahesh, K. Liu,R. C. Panicker and S. Q. Yao, Chem. Commun., 2007, 1518-1520. J. J.Wang, X. Feng, Z. Xun, D. Q. Shi and Z. B. Huang, J. Org. Chem., 2015,80, 8435-42. Forristal, I. J. Sulphur Chem., 2005, 26, 163. F. Hof, A.Schutz, C. Fah, S. Meyer, D. Bur, J. Liu, D. E. Goldberg, and E.Diederich, Angew. Chem., Int. Ed., 2006, 45, 2138-2141]. The compoundscontaining sulfone group is very important in medicinal chemistry suchas Rofecoxib, non-steroidal anti-inflammatory (NSAID) used in thetreatment of arthritis and other similar conditions causing chronic oracute pain, Sulindac sulfone is used in the treatment of tumor and HCA-7cells which leads to inhibition of prostaglandin E2 production [C. S.Williams, A. P. Goldman, H. Sheng, J. D. Morrow, and R. N. D. Bois,Neoplasia., 1999, 1, 170-176] and Bicalutamide used in the treatment ofprostate cancer [P. F. Schnellhammer, Expert Opin. Pharmacother, 2002,3, 1313-1328]. Other sulfone derivatives have been used as drugs due totheir high potential as antibacterial, antifungal, anti-nociceptive andanti-inflammatory agents such ad Lasix, Aquazide h, Sulfadimidine [K.Rais-Bahrami, M. Majd, E. Veszelovszky, and B. L. Short, Am. J.Perinatol., 2004, 21, 329-332. J. D. Duarte, and R. M. Cooper-DeHoff,Expert Rev Cardiovasc Ther., 2010, 8, 793-802. A. Romvary, and F. Simon,Acta Vet Hung., 1992, 40, 99-106. W. Billard, H. Binch, K. Bratzler, L.-Y., Chen, G. C. Jr, R. A. Duffy, S. Dugar, J. Lachowicz, R., McQuade,P. Pushpavanam, V. B. Ruperto, L. A. Taylor, and J. W. Clader, Bioorg.Med. Chem. Lett. 2000, 10, 2209-2212. V. Padmavathi, P. Thriveni, G. S.Reddy, and D. Deepti, Eur. J. Med. Chem., 2008, 43, 917-924. N. K.Konduru, S. Dey, M. Sajid, M. Owais, and N. Ahmed,. Eur. J. Med. Chem.,2013 59, 23-30. P. A. Todd, and S. P. Clissold, Tenoxicam. Drugs., 1991,41, 625-646. C. R. Lee, and J. A. Balfour, (1994). Piroxicam-βCyclodextrin. Drugs., 1994, 48, 907-929.] well-known drugs in marketDapsone and Promine [G. H. Faget, R. C. Pogge, F. A. Johansen, J. F.Dinan, B. M. Prejean, and C. G. Eccles, Int J Lepr Other Mycobact Dis.,1966, 34, 298-310.] The sulfone moiety also finds its value in theagrochemical field such as Mesotrione and Cafenstrole reported asherbicide [G. Mitchell, D. W. Bartlett, T. E. M. Fraser, T. R. Hawkes,D. C. Holt, J. K. Townson, and R. A. Wichert, Pest Manag Sci., 2001, 57,120-128. P. Boger. J Pest Sci., 2003, 28, 324].

Background of PI3K and MK2:

The phosphoinositide 3-kinase (PI3K) is one of the most studied kinases,which plays pivotal role in regulation of various cellular processes andpathogenesis of Inflammation, Cancer and Diabetes. Kinases of PI3Kfamily acts separately as well as in synergistic manner to regulatecrucial signaling and metabolic processes. There is an augmentedrecognition of the key importance of three different PI3K classes (I, IIand III) with their isoforms in various disease progression [S. Jean,and A. A. Kiger, 2014].

Due to their key crosstalk between themselves and kinases form othersignaling pathways, class I PI3K isoforms have always been epicenter ofthe drug discovery as potent drug targets in the management of variousdiseases and helps in the quest of class-I-related therapies [A.Carracedo, and P. P. Pandolfi, Oncogene., 2008, 27, 5527].

Role of PI3K:

Inflammation: Previous studies have found that class I PI3K isoformshave great significance in inflammation and immunomodulation [A. K.Stark, S. Sriskantharajah, E. M. Hessel, and K. Okkenhaug, Currentopinion in pharmacology, 2015, 23, 82-91] [D. A. Fruman, and C. Rommel,Nature reviews Drug discovery, 2014, 13, 14]. Ubiquitous presence ofPI3K-p110α is essential for angiogenesis and insulin signaling.PI3K-p110/120γ and PI3K-p110δ are mostly cell surface receptor dependentand are found in different immune cells like macrophages, dendriticcells, mast cell, T cells and B cell regulating growth anddifferentiations through signal transmissions.

Since numerous studies have established the fact that PI3K kinase isinvolved in key signalling and metabolic pathways of inflammation,rheumatoid arthritis and senescence, therefore, considering the factsthat in diseases where pro-inflammatory cytokine response isdysregulated, PI3K kinases could be a promising therapeutic target.

Cancer: Numerous studies linked the phosphoinositide 3-kinase (PI3K)pathway with activation of receptor tyrosine kinases (RTKs), Gprotein-coupled receptors (GPCRs) and oncogenes such as RAS whichultimately affects various key cellular functions including cellproliferation, apoptosis and tumorigenesis. Along with it, comprehensivegenome analysis of different human tumor samples have revealed thepresence of mutations or alterations in components of PI3K signallingpathways [R. K. Thomas, A. C. Baker, R. M. DeBiasi, W. Winckler, T.LaFramboise, W. M. Lin, M. Wang, W. Feng, T. Zander, L. E. MacConaill,and J. C. Lee, Nat Genet., 2007, 39, 347; Y. Samuels, Z. Wang, A.Bardelli, N. Silliman, J. Ptak, S. Szabo, H. Yan, A. Gazdar, S. M.Powell, G. J. Riggins, and J. K. Willson, Science., 2004, 304, 554-554].Inclusively, PI3K has been directly implicated in number of cancersincluding colon, brain, gastric, breast and lung cancer. [L. D. Wood, D.W. Parsons, S. Jones, J. Lin, T. Sjöblom, R. J. Leary, D. Shen, S. M.Boca, T. Barber, J. Ptak, and N. Silliman, Science., 2007, 318,1108-1113; R. K. Thomas, A. C. Baker, R. M. DeBiasi, W. Winckler, T.LaFramboise, W. M. Lin, M. Wang, W. Feng, T. Zander, L. E. MacConaill,and J. C. Lee, Nat Genet., 2007, 39, 347; Cancer Genome Atlas ResearchNetwork, Nature., 2008, 455, 1061; L. Ding, G. Getz, D. A. Wheeler, E.R. Mardis, M. D. McLellan, K. Cibulskis, C. Sougnez, H. Greulich, D. M.Muzny, M. B. Morgan, and L. Fulton, Nature., 2008, 455, 1069; Y.Samuels, Z. Wang, A. Bardelli, N. Silliman, J. Ptak, S. Szabo, H. Yan,A. Gazdar, S. M. Powell, G. J. Riggins, and J. K. Willson, Science.,2004, 304, 554-554; A. Di Cristofano, M, De Acetis, A. Koff, C.Cordon-Cardo, and P. P. Pandolfi, Nat Genet., 2001, 27, 222;, D. W.Parsons, S. Jones, X. Zhang, J. C. H. Lin, R. J. Leary, P. Angenendt, P.Mankoo, H. Carter, I. M. Siu, G. L. Gallia, and A. Olivi, Science.,2008, 321, 1807-1812; P. Liu, H. Cheng, T. M. Roberts, and J. J. Zhao,Nat Rev Drug Discov., 2009, 8, 627].

Rheumatoid arthritis: Recent studies had shown that targeting PI3Kisoforms in a mouse model of systemic lupus erythematosus can reduceglomerulonephritis and suppresses disease progression in rheumatoidarthritis mouse models [C. Rommel, M. Camps and, H. Ji, Nature ReviewsImmunology., 2007, 7, 191].

Lung fibrosis and Asthma: Regulation of inflammation and fibrosis hasbeen shown to be governed by four class I PI3Ks [C. C. Campa, R. L.Silva, J. P. Margaria, T. Pirali, M. S. Mattos, L. R. Kraemer, D. C.Reis, G. Grosa, F. Copperi, E. M. Dalmarco, and R. C. Lima-Júnior,.Nature communications., 2018, 9, 5232]. For example, a variety ofleukocyte functions, including proliferation, differentiation,migration, and survival, are controlled by PI3Kγ and δ. Furthermore,proliferation of different lung cell types is governed by PI3Kα and β,PI3Kγ and β has been found involved in pulmonary inflammation andfibrotic remodelling which ultimately leads to lung fibrosis and asthma.[P. T. Hawkins, and L. R. Stephens, PI3K signalling in inflammation,Biochimica et Biophysica Acta (BBA)-Molecular and Cell Biology ofLipids, 2015, 1851, 882-897; A. Ghigo, F. Damilano, L. E. Bioessays, 32,185-196, R. C. E. Roffe, A. L. Souza, L. P. Sousa, M. Mirolo, A. Doni,and G. D, J. Leukoc. Biol., 2011, 89, 269-282]. Diabetes relateddisorders: PI3K regulates glucose uptake in the cells by downstreamactivation of AKT and hexokinase. Increase in glucose uptake is theimmediate effect of insulin driven PI3K signalling in muscle and fatcells, attributable to increased glucose translocation to the membraneand upregulation of the genes encoding the transporters [[L. C. Cantley,and Z. Songyang, J Cell Sci, 1994, 121-126]. Receptor tyrosine kinasesundergo conformational changes after stimulation of the growth factor,allowing them to auto-phosphorylate and become active. Subsequently,AKT2 is phosphorylated and inhibits RabGAP and AS160, resulting intomigration of glucose transporter GLUT4 to the plasma membrane [J.Yuasa-Kawada, M. Kinoshita-Kawada, Y. Rao, and J. Y. Wu, Proceedings ofthe National Academy of Sciences, 2009, 106, 14530-14535; L. C.Cantley,. Science, 2002, 296, 1655-1657]

Mapkapk2

MAPKPAK2 or, MK2 is downstream substrate of p-38 MAPK in MAPK pathwayand have been found to be extremely involved in oxidative stress,inflammation, regulation of cell cycle, tumorigenesis and cell migration[Gurgis, Fadi Maged Shokry, William Ziaziaris, and Lenka Munoz, MolPharmacol., 2014, 345-356; Maruyama, Junichi, et al. Curr Med Chem.,2009, 1229-1235; Jackson, Robert M., and Rolando Garcia-Rojas, Exp LungRes., 2008, 245-262]. Key studies elucidated its role at posttranscriptional level in gene expression, cell proliferation andapoptosis through modulation of transcript stability via RBPs (RNABinding Proteins) [Pearson, Gray, et al. Endocr Rev., 2001, 153-183; S.Soni, P. Anand, and Y. S. Padwad, J Exp Clin Cancer Res., 2019, 38, 121;S. Soni, M. K. Saroch, B. Chander, N. V. Tirpude, and Y. S. Padwad, JExp Clin Cancer Res., 2019, 38, 175]. Furthermore, MK2 has been found toregulate various cellular processes in response to extracellular signalsincluding oxidative stress, inflammation, infection, radiation andgenotoxicity [S. J. Diaz-Cano, Re. Pomerance et al. J. Pathol., 2006,209, 298-306; J. Pathol., 2006, 210, 133, A. Kotlyarov, Y. Yannoni, S.Fritz, K. LaaB, J. B. Telliez, D. Pitman, L. L. Lin and M. Gaestel, MolCellular Biol. 2002, 22, 4827-35].

Upon activation from p38-MAPK, cytoplasmic localization of MK2 takesplace with the help of nuclear export signals (NES) [Pearson, Gray, etal. Endocrine reviews, 2001, 153-183;, S. Soni, P. Anand, and Y. S.Padwad, Journal of Experimental & Clinical Cancer Research., 2019, 38,121,; S. J. Diaz-Cano, Re. Pomerance et al. J. Pathol, 2006, 209,298-306. J. Pathol., 2006, 210, 133], where it directly acts upon itscrucial downstream substrates like Hsp27 (Heat Shock protein 27) andleads to remodeling of cellular cytoskeleton, cell migration, cellinvasion and metastasis [F. M. Gurgis, W, Ziaziaris and L. Munoz, MolPharmacol. 2014, 85, 345-356, Rogalla, Thorsten, et al. J Biol Chem.,1999, 18947-18956].]. Additionally, MK2 favors the cell survival byreversing post chemotherapy DNA damage through activating G2/M arrestvia checkpoint signaling, ultimately imposing resistance to thetherapeutic protocol [Cannell, Ian G., et al. Cancer cell., 2015,623-637].

Role of MK2:

Inflammation:

MK2 extensively regulates the biosynthesis of TNFα and production ofpro-inflammatory mediators like TNFα, IL-1, IL-β, IL-6, interferon-γ(IFNγ) and other cytokines [Rogalla, Thorsten, et al. J Biol Chem.,1999, 18947-18956]. MK2 proved to be essential for the stimulation ofcytokine biosynthesis through LPS-induced upregulation of cytokine mRNAstability and translation. This is supported by a substantial reductionin TNFα synthesis despite of LPS induction in MK2 deficient transgenicmice model [R. Winzen, M. Kracht, B. Ritter, A. Wilhelm, C. Y. A. Chen,A. B. Shyu, M. Müller, M. Gaestel, K. Resch, and H. Holtmann, The EMBOjournal., 1999, 18, 4969-4980; Rogalla, Thorsten, et al. J Biol Chem.,1999, 18947-18956]. MK2 has also been implicated in RIPK1 inhibition bylimiting its cytotoxic and apoptotic activity, hence, increase cytokineexpression during inflammation [M. B. Menon, J. GropengieBer, J.Fischer, L. Novikova, A. Deuretzbacher, J. Lafera, H. Schimmeck, N.Czymmeck, N. Ronkina, A. Kotlyarov, and M. Aepfelbacher, Nat Cell Biol.,2017, 19, 1248].

Cancer: A vast array of studies has been reported direct role of MK2 incell cycle regulation, modulation of RBPs, uncontrolled cellproliferation, tumorigenesis and metastasis. In the events of cellcycle, MK2 phosphorylates CDC25 family members (CDC25B and CDC25C) topersuade 14-3-3 binding which leads to the arrest of cell cycle [I. A.Manke, A. Nguyen, D. Lim, M. Q. Stewart, A. E. Elia, and M. B. Yaffe,Mol Cell., 2005, 17, 37-48]. MK2 also targets the activation ofubiquitin ligase HDM2 to promote degradation of p53 leading to post DNAdamage cell survival [H. O. Weber, R. L. Ludwig, D. Morrison, A.Kotlyarov, M. Gaestel, and K. H. Vousden, Oncogene., 2005, 24, 1965] andhas also been linked with imposing resistance to the apoptosis caused byp53 mutation [F. M. Gurgis, W, Ziaziaris and L. Munoz, Mol Pharmacol.2014, 85, 345-56].

Additionally, MK2 plays role in Mdm2 activation, resulting into Mdm2mediated p53 inactivation and degradation, confirmed by rise in p53level in MK^(−/−) cells in contrast to low Mdm2 levels [H. O. Weber, R.L. Ludwig, D. Morrison, A. Kotlyarov, M. Gaestel, and K. H. Vousden,Oncogene., 2005, 24, 1965]. These studies have established MK2 as a DNAdamage checkpoint kinase functioning parallel to conventional CHK1 andCHK2 [Cannell, Ian G., et al. Cancer cell., 2015, 623-637, I. A. Manke,A. Nguyen, D. Lim, M. Q. Stewart, A. E. Elia, and M. B. Yaffe, Molecularcell, 2005, 17, 37-48, M. B. Menon, J. GropengieBer, J. Fischer, L.Novikova, A. Deuretzbacher, J. Lafera, H. Schimmeck, N. Czymmeck, N.Ronkina, A. Kotlyarov, and M. Aepfelbacher, Nat Cell Biol., 2017 19,1248]. MK2 orchestrate mRNA stability of genes as well asphosphorylation and expression of numbers of proteins involved in immuneresponse [J. S. Erdem, V. Skaug, A. Haugen and S. Zienolddiny, JCancer., 2016, 7, 512], cell cycle [[F. M. Gurgis, W, Ziaziaris and L.Munoz, Mol Pharmacol. 2014, 85, 345-56.], cytoskeleton remodelling[Rogalla, Thorsten, et al. J Biol Chem., 1999, 18947-18956; A.Kotlyarov, Y. Yannoni, S. Fritz, K. LaaB, J. B. Telliez, D. Pitman, L.L. Lin and M. Gaestel, Mol Cellular Biol., 2002, 22, 4827-35.50; 51. B.Kumar, S. Koul, J. Petersen, L. Khandrika, J. S. Hwa, R. B. Meacham, S.Wilson and H. K. Koul, Cancer Res., 2010, 70, 832-41.51], evasion ofapoptosis and cell migration [Rogalla, Thorsten, et al. J Biol Chem.,1999, 18947-18956 40; F. M. Gurgis, W, Ziaziaris and L. Munoz, MolPharmacol. 2014, 85, 345-56; B. Kumar, S. Koul, J. Petersen, L.Khandrika, J. S. Hwa, R. B. Meacham, S. Wilson and H. K. Koul, CancerRes., 2010, 70, 832-41.] in response to external stimuli. These stimuliare abundant in tumor microenvironment and most of the time initiate theactivation of MK2 and subsequently its downstream substrates like Hsp27,modulating the levels of cytokines, chemokines and matrixmetalloproteinases (MMPs), finally resulting into facilitation of tumormicroenvironment, neoangiogenesis and extravasation.

Various key studies performed in the area specifically support thedirect involvement of MK2 and its downstream substrate Hsp27 in thepathogenesis of different cancers, prominently including intestinalcancer [A. Henriques, V. Koliaraki, G. Kollias, and Mesenchymal, ProcNatl Acad Sci., USA. 2018, 115, E5546-55,; H. R. Cheruku., A.Mohamedali, D. I. Cantor, S. H. Tan, E. C. M. S. Nice. EuPA OpenProteom., 2015, 8, 104-15.], skin cancer [C. Johansen, C. Vestergaard,K. Kragballe, G. Kollias, M. Gaestel, and L. Iversen, Carcinogenesis.,2009, 30, 2100-8], bladder cancer [B. Kumar, J. Sinclair, L. Khandrika,S. Koul, S. Wilson and H. K. Koul, Int J Oncol., 2009, 34, 1557-64.],prostate cancer [S. A. Hayes, X. Huang, S. Kambhampati, L. C. Plataniasand R. C. Bergan, Oncogene., 2003, 22, 4841.], lung cancer [B. Liu, L.Yang, B. Huang, M. Cheng, H. Wang, Y. Li, D. Huang, J. Zheng, Q. Li, X.Zhang, and W. Ji, Am J Hum Genet., 2012, 91, 384-390, J. S. Erdem, V.Skaug, A. Haugen, and S. Zienolddiny, J. Cancer., 2016, 7, 512] and headand neck cancer [S. Soni, M. K. Saroch, B. Chander, N. V. Tirpude, andY. S. Padwad, J Exp Clin Cancer Res. 2019, 38, 17535].

Alzheimer's disease and Neuroinflammation: Current evidences supportsthe role of MK2 in regulation of various inflammatory cytokinesexpressions in neuroinflammation-associated important brain disorders.For example, MK2 was found to be overexpressed in LPS⁺ IFF-gammastimulated microglial cells. Ex-vivo cultured microglia from MK2^((−/−))mice have shown significant inhibition in the release of TNFα and MIP-1α(macrophage inflammatory protein-1α) as compared to MK2^((+/+))wild-type microglia. In addition to this, deficiency of MK2 in APP-PS1transgenic Alzheimer mouse promotes autophagy/macroautophagy. Based onthe evidences from various studies, it is clear that MK2 contributes tothe pathophysiology of Alzheimer's disease and Parkinson's disease [F.M. Gurgis, W, Ziaziaris and L. Munoz, Mol Pharmacol. 2014, 85, 345-56,A. A. Culbert, S. D. Skaper, D. R. Howlett, N. A. Evans, L. Facci, P. E.Soden, Z. M. Seymour, F. Guillot, M. Gaestel, and J. C. Richardson, JBiol Chem., 2006, 281, 23658-23667, S. A. Correa, and K. L. Eales,Journal of signal transduction., 2012, J. Alam, and W. Scheper,Autophagy., 2016, 12, 2516-2520.].

Lung injury and pulmonary fibrosis: MK2 has been long implicated in theinflammation via activation of NADPH oxidase, regulation of TNF-αlevels, neutrophil recruitment and cell cycle arrest. These mechanismshave been found to be involved in the molecular pathogenesis of acutelung injury, pulmonary fibrosis, and non-small-cell lung cancer [F.Qian, J. Deng, G., Wang, D Ye, R. and W Christman, Curr Protein PeptSci., 2016, 17, 332-342]. Increase in MK2 mediated inflammatorycytokines mRNAs results in actin reorganization, change in cell adhesionproperties, increase in α-SMA (α-smooth muscle actin) protein expressionand myofibroblast differentiation. Combinatory, all these events lead toregulation of cytoskeleton structure and facilitates lung injury andpulmonary fibrosis [L. B. Lopes, E. J. Furnish, P. Komalavilas, C. RFlynn, P. Ashby, A. Hansen, D. P. Ly, G. P. Yang, M. T. Longaker, A.Panitch, and C. M. Brophy, J Invest Dermatol., 2009, 129, 590-598.61; R.K. Singh and A. K. Najmi, Curr Drug Targets., 2019, 20, 367-379, R.Vittal, A. Fisher, H. Gu, E. A. Mickler, A. Panitch, C. Lander, O. W.Cummings, G. E. Sandusky, and D. S. Wilkes, Am J Respir Cell Mol Biol.,2013, 49, 47-57.].

Asthma: It has been observed in the studies that, MK2 is necessary toproduce localized Th2-type inflammation and subsequently developsexperimental asthma. Although MK2 doesn't affect systemic Th2 immunity,but it reduces expression of chemokines, adhesion molecules and alsodecreases endothelial permeability. Additionally, transcriptionalexpression of chemokines and adhesion molecules have been found toassociate with NF-κB and it is well versed that MK2 and its downstreamHsp27 are essential for sustained production of NF-κB [M. M. Gorska, Q.Liang, S. J. Stafford, N. Goplen, N. Dharajiya, L. Guo, S. Sur, M.Gaestel, and R. Alam, J. Exp. Med., 2007, 204, 1637-1652].

Rheumatoid arthritis: TNF-alpha has been shown to play an important rolein Rheumatoid arthritis (RA) by mediating immune regulation andinflammatory response. MK2 has been long associated with biosynthesis ofTNF-alpha at a posttranscriptional level, so the studies based on thissupports the direct involvement of MK2 in RA. This has been confirmed inthe DBA/1Lacj collagen-induced arthritis (CIA) mice model by deletion ofthe MK2 gene, which enhanced protection against CIA [M. Hegen, M.Gaestel, C. L. Nickerson-Nutter, L. L. Lin, and J. B. Telliez, J.Immunol., 2006, 177, 1913-1917].

Atherosclerosis: Study has been reported the functional role of MK2 andatherogenesis in hypercholesterolemia. Activated form of MK2 has beendetected in endothelium and macrophage-rich plaque areas inside aortasof hypercholesterolemic LDL receptor deficient mice. Contrary, deletionof MK2 in hypercholesterolemic 1dlr(−/−) mice (1dlr(−/−)/mk2(−/−))decreases the accumulation of lipid and macrophages in the aorta despiteof an artherogenic diet and increased plasma lipoprotein levels [K.Jagavelu, U. J. Tietge, M. Gaestel, H. Drexler, B. Schieffer, and U.Bavendiek, Circ Res., 2007, 101, 1104-1112].

Therefore, there is a need in the art for novel compounds for treatmentof PI3K and MK2 related disorders.

Objectives of the Invention

The main objective of the present invention is to provide sulfonamidefused benzocycloheptenes of general formula I useful for treatment ofdiseases responsible through PI3K and MK2 pathway.

Yet another objective of the present invention is to provide furan andpyrrol substituted benzocycloheptenes of general formula II useful asPI3K and MK2 inhibitor for treatment of diseases.

Still another objective of the present invention is to provide alkylcarbonyl substituted benzocycloheptenes of general formula III useful asPI3K and MK2 inhibitor, for treatment of diseases.

Yet another objective of the present invention is to provide multi-ringfused quinolinone and quinolinol substituted benzocycloheptenes ofgeneral formula IV and V useful as PI3K and MK2 inhibitor, for treatmentof diseases.

Still another objective of the present invention is to provide sulfonesubstituted benxocycloheptenes of general formula VI useful as PI3K andMK2 inhibitor, for treatment of diseases.

Yet another objective of the present invention is to provide a methodfor preparation of benzocycloheptene analogues of formula I-VI fromhimachalenes collected from Cedrus deodara oil.

SUMMARY OF THE INVENTION

As aspect of present invention provides a compound of general formula I,II, III, IV, V or VI:

and pharmaceutical acceptable salts and enantiomers thereof,

-   -   wherein Y is selected from the group consisting of C, O, N and        S;    -   Z is selected from the group consisting of H, C1-C6 alkyl, C1-C6        aryl, halogen, oxygen, nitrogen and sulfur;    -   R¹ is selected from the group consisting of hydrogen, hydroxyl,        nitrogen substituted group selected from benzylamine or aniline,        S-containing group selected from thiophene, carboxylic acid and        its derivatives selected from propanoic acid, and halogen;    -   R² is selected from the group consisting of H, C1-C6 alkyl,        C1-C6 aryl, halide, C1-C6 amines and C1-C6 carbonyl group;    -   R³ is selected from the group consisting of H, C1-C6 alkyl,        C1-C6 aryl and sulfur containing group selected from thiophene;    -   X is selected from the group consisting of carbon, nitrogen,        oxygen and sulfur;    -   R⁴ is selected from the group consisting of H, C1-C6 alkyl,        C1-C6 aryl, halogen and carbonyl group selected from aldehyde        substituted benzenesulphonamide;    -   R⁵ is selected from the group consisting of H, C1-C6 alkyl,        C1-C6 aryl, halogen and C1-C6 carbonyl group;    -   R⁶ is selected from the group consisting of H, C1-C6 alkyl,        C1-C6 aryl and C1-C6 amine group.

Another aspect of the present invention provides a compound of formulaI-VI for use as anti-type 2 diabetes, antipyretic, anti-inflammatory,anticancer, antiulcer, CNS-stimulant, and CNS-depressant.

Yet another aspect of the present invention provides a pharmaceuticalformulation comprising the compound of formula I-VI along with apharmaceutically acceptable adjuvant, diluent or carrier.

Still another aspect of the present invention provides a compound offormula I-VI, for use as an inhibitor of PI3K and MK2 mediated activity.

BRIEF DESCRIPTION OF DRAWINGS

FIG. 1 : Image depicts the on-rate and off-rate of binding of evaluatedcompounds of general formula II to PI3K (120y). Plotting associationrate constant ka against dissociation rate constant kd (here onlogarithmic scales) creates a plot where the affinity is represented bydiagonal lines. Compounds on the same diagonal have the same affinitybut differ in kinetics.

FIG. 2 : Graphical representation of the on-rate and off-rate of bindingof evaluated compounds of general formula II to PI3K (120y).

FIG. 3 : Graphs represent the dose dependent inhibition of the PI3K(120y) by compounds of general formula II (5, 7, 8, 9, 12 and 16).

FIG. 4 : Image depicts the on-rate and off-rate of binding of evaluatedcompounds of general formula II to MK2. Plotting association rateconstant ka against dissociation rate constant kd (here on logarithmicscales) creates a plot where the affinity is represented by diagonallines. Compounds on the same diagonal have the same affinity but differin kinetics.

FIG. 5 : Graphical representation of the on-rate and off-rate of bindingof evaluated compounds of general formula II to MK2

FIG. 6 : Graphs represent the dose dependent inhibition of the MK2 bycompounds of general formula II (1, 3, 6, 8, 12, 13, 14, 15 and 17 alongwith marketed MK2-inhibitor PF 3644022.

DETAILED DESCRIPTION OF THE INVENTION

The present invention is directed towards substituted benzocyclohepteneanalogues, of general formula I, II, III, IV, V or VI

wherein

-   -   in Formula I-VI,    -   R¹ is selected from the group consisting of H, OH, N substituted        group selected from benzylamine or aniline, S-containing group        selected from thiophene, carboxylic acid and its derivatives        selected from propanoic acid, and halogen;    -   R² is selected from the group consisting of H, C1-C6 alkyl,        C1-C6 aryl, halide, C1-C6 amine and C1-C6 carbonyl group;    -   in Formula I, R³—R⁵ are each independently selected from the        group consisting of H, C1-C6 alkyl, C1-C6 aryl, halogen and        C1-C6 carbonyl group;    -   in Formula II, Y is selected from the group consisting of C, O,        N and S;    -   in Formula III, Z is selected from the group consisting of H,        C1-C6 alkyl, C1-C6 aryl, halogen, oxygen, nitrogen and sulfur;    -   in Formula IV, n varies from 5 to 6;    -   in Formula V, n varies from 5 to 6 and R³ is selected from the        group consisting of H, OH, N substituted group selected from        pyridine, carboxylic acid and its derivatives selected from        propanoic acid and halogen;    -   in Formula VI, R⁵ is selected from the group consisting of H,        halide, oxygen, nitrogen containing group selected from the        group consisting of enaminone, cyclohexane 1,3-dione, C1-C6        alkyl and C1-C6 aryl group; and R⁶ is selected from the group        consisting of C1-C6 alkyl, C1-C6 aryl, hydrogen and hetero atoms        selected from thiophene.

The present invention provides a compound of general formula I, II, III,IV, V or VI:

and pharmaceutical acceptable salts and enantiomers thereof,

-   -   wherein    -   Y is selected from the group consisting of C, O, N and S;    -   Z is selected from the group consisting of H, C1-C6 alkyl, C1-C6        aryl, halogen, oxygen, nitrogen and sulfur;    -   R¹ is selected from the group consisting of hydrogen, hydroxyl,        nitrogen substituted group selected from benzylamine or aniline,        S-containing groups selected from thiophene, carboxylic acid and        its derivatives selected from propanoic acid, and halogen;    -   R² is selected from the group consisting of H, C1-C6 alkyl,        C1-C6 aryl, halide, C1-C6 amine and C1-C6 carbonyl groups;    -   R³ is selected from the group consisting of H, C1-C6 alkyl,        C1-C6 aryl and sulfur containing group selected from thiophene;    -   X is selected from the group consisting of carbon, nitrogen,        oxygen and sulfur;    -   R⁴ is selected from the group consisting of H, C1-C6 alkyl,        C1-C6 aryl, halogen and carbonyl group selected from aldehyde        substituted benzenesulphonamide;    -   R⁵ is selected from the group consisting of H, C1-C6 alkyl,        C1-C6 aryl, halogen and C1-C6 carbonyl group; and    -   R⁶ is selected from the group consisting of H, C1-C6 alkyl,        C1-C6 aryl and C1-C6 amine group.    -   In an embodiment of the present invention there is provided a        compound of the formula (I),    -   wherein R¹ is selected from the group consisting of H, OH, N        substituted group selected from benzylamine or aniline,        carboxylic acid and its derivatives selected from propanoic acid        and halogens,    -   R² is selected from the group consisting of H, C1-C6 alkyl,        C1-C6 aryl, halide, C1-C6 amine and C1-C6 carbonyl group; and    -   R³-R⁵ are each independently selected from the group consisting        of H, C1-C6 alkyl, C1-C6 aryl, halogen and C1-C6 carbonyl group.    -   In another embodiment of the present invention there is provided        a compound of the formula (II),    -   wherein R¹ is selected from the group consisting of H, OH, N        substituted group selected from benzylamine, or aniline,        carboxylic acid and its derivatives selected from propanoic        acid, and halogen,    -   R² is selected from the group consisting of H, C1-C6 alkyl,        C1-C6 aryl, halide, C1-C6 amine and C1-C6 carbonyl group; and    -   and Y is selected from the group consisting of C, O, N and S.    -   In yet another embodiment of the present invention there is        provided a compound of the formula (III),    -   wherein R¹ is selected from the group consisting of H, OH, N        substituted group selected from benzylamine or aniline,        carboxylic acid and its derivatives selected from propanoic        acid, and halogen,    -   R² is selected from the group consisting of H, C1-C6 alkyl,        C1-C6 aryl, halide, C1-C6 amine and C1-C6 carbonyl group; and    -   Z is selected from the group consisting of H, C1-C6 alkyl, C1-C6        aryl, halogen, oxygen, nitrogen and sulfur group.    -   In still another embodiment of the present invention there is        provided a compound of the formula (IV),    -   wherein R¹ is selected from the group consisting of H, OH, N        substituted group selected from benzylamine, or aniline,        carboxylic acid and its derivatives selected from propanoic acid        and halogen,    -   R² is selected from the group consisting of H, C1-C6 alkyl,        C1-C6 aryl, halide, C1-C6 amine, and C1-C6 carbonyl group; and        and n varies from 5 to 6.    -   In another embodiment of the present invention there is provided        a compound of the formula (V),    -   wherein R¹ is selected from the group consisting of H, OH, N        substituted group selected from benzylamine or aniline,        carboxylic acid and its derivative selected from propanoic acid        and halogen,    -   R² is selected from the group consisting of H, C1-C6 alkyl,        C1-C6 aryl, halide, C1-C6 amine, and C1-C6 carbonyl group;    -   n varies from 5 to 6; and    -   R³ is selected from the group consisting of H, CJ-C6 alkyl,        CJ-C6 aryl and halogen group.    -   In yet another embodiment of the present invention there is        provided a compound of the formula (VI),    -   wherein R¹ is selected from the group consisting of H, OH, N        substituted group selected from benzylamine, or aniline,        carboxylic acid and its derivatives selected from propanoic acid        and halogen;    -   R² is selected from the group consisting of H, CJ-C6 alkyl,        CJ-C6 aryl, halide, CJ-C6 amine and CJ-C6 carbonyl group;    -   R⁵ is selected from the group consisting of hydrogen, CJ-C6        alkyl, halide, sulphur, C1-C6 amine and CJ-C6 aryl group; and    -   R⁶ is selected from the group consisting of H, C1-C6 alkyl,        C1-C6 aryl and C1-C6 amine group.

In still another embodiment of the present invention there is provided acompound of the formula I-VI, for use as anti-type 2 diabetes,antipyretic, anti-inflammatory, anticancer, antiulcer, CNS-stimulant,and CNS-depressant.

Another embodiment of the present invention provides a pharmaceuticalformulation comprising the compound of formula I-VI along with apharmaceutically acceptable adjuvant, diluent or carrier.

In yet another embodiment of the present invention there is provided acompound of the formula I-VI, for use as an inhibitor of PI3K and MK2mediated activity.

These substituted benzocycloheptene analogues are semi-synthesized fromhimachalenes extracted from Cedrus deodara oil.

The present invention discloses synthesis of benzocycloheptene compoundsusing easily available and low cost natural precursors i.e. Cedrusdeodara oil following less steps, economic and cost-effective approach.

Under this investigation, semi-synthetic approaches have been developedto minimize number of challenging steps in organic synthesis andavoiding high cost reagents and chemicals to reduce overall cost ofproduction.

Under this investigation, different new methodologies have beendeveloped to overcome the existing challenges in this type of organicsynthesis. High purity of the molecules (>98-99%) has been achievedthrough column chromatography.

The process is easy to apply for scale-up synthesis which is a veryimportant issue to evaluate biological activities and future industrialinterest.

These class of compounds are known for different biological activitiestherefore, facile approaches have been developed for different newclasses of benzocycloheptene analogues synthesis following new processand further applied for treatment of PI3K and MK2 related disorders.

Further, the use of natural analogues also reduces the toxic effect andits specific structure enhances the chance of biological activities fortherapeutic development.

Compounds constituted of complex bicyclic framework with alkyl group aredifficult to introduce. Thus, the core structure is derived from naturalprecursor which reduces the cost of production, reagents and overallmakes the process economic.

Compounds constituted from natural precursor are less toxic in natureand indicated different applications for treatment of PI3K and MK2related disorders responsible for different diseases.

EXAMPLES

The following examples are given by way of illustration and thereforeshould not construed to limit the scope of the present invention.

Experimental Part

All reagents and solvents were purchased from commercial sources(Sigma-Aldrich, Merck India Ltd). Reactions were monitored by TLC platescoated with 0.2 mm silica gel 60 F₂₅₄. TLC plates were visualized by UVirradiation (254 nm) and iodine spray. The products were purified bycolumn chromatography employing silica gel of 60-120 mesh size (Merck).The ¹H and ¹³C NMR spectra were recorded at 298 K with a Bruker AM-300spectrometer; using TMS as internal reference standard in CDCl₃. HRMSwere conducted with UHR-QTOF (ultra-high resolution Q-time of flight).IR spectra were obtained on a Nicolet 5700 FTIR (Thermo, USA)spectrophotometer in the region 4,000-400 cm-1 using KBr disks. CEMDiscover™ focused microwaves (2450 MHz, 300 W) were used. Thetemperature on the surface of the reaction flask was measured with aninbuilt infrared temperature probe in the microwave experiment. Thecoupling constants (J) are reported in hertz (Hz) and the followingabbreviations are used to designate signal multiplicity: s=singlet;d=doublet; t=triplet; m=multiplet; br=broad singlet.

Example 1

General Procedure for the Synthesis of Formula I

Synthesis ofN-benzyl-N-((8-bromo-3,5,5-trimethyl-6,7-dihydro-5H-benzo[7]annulen-9-yl)methyl)-4-methylbenzenesulfonamide

A mixture of8-bromo-9-(bromomethyl)-3,5,5-trimethyl-6,7-dihydro-5H-benzo[7]annulene(0.279 mmol, 1.0 equiv.), N-benzyl-4-methylbenzzenesulfonamide (0.41mmol, 1.5 equiv.), K₂CO₃ (0.55 mmol, 2 equvi.) in DMF (3 ml) were placedin reaction tube (15 mL) at 90° C. for 16 h. After cooling the reactionmixture to ambient temperature, water was added and extracted with ethylacetate (10×3 times). The combined organic layer was washed with waterand dried over Na₂SO₄ and vacuum evaporated. The viscous liquid obtainedwas purified by column chromatography on silica gel (mesh 60-120) usingthe mixture of hexane: EtOAc (80:20) to obtain the desired product asyellow oily (45 mg, 30%).

¹H NMR (CDCl₃, 300 MHz, δ, ppm): 1.08 (s, 6H), 1.94-1.89 (m, 2H),2.32-2.24 (m, 8H), 4.33 (s, 2H), 4.36 (s, 2H), 6.96-6.94 (d, J=7.8, 1H),7.08-7.05 (d, 3H), 7.21-7.11 (m, 5H), 7.36-7.29 (m, 3H); ¹³C NMR (CDCl₃,75 MHz) δ, 21.4, 21.5, 31.6, 37.6, 38.6, 47.3, 51.8, 52.6, 126.6, 126.6,127.2, 127.3, 128.1, 129.0, 133.3, 134.5, 136.4, 136.9, 137.1, 142.8,146.2. ESI-MS: calcd for C₂₉H₃₂BrNO₂S [M+H]⁺ 538.1410, found 538.0189.

Example 2

General Procedure for the Synthesis of Formula II

Synthesis of6,6,8-trimethyl-1,4,5,6-tetrahydro-3H-benzo[3,4]cycloheptal[1,2-c]furan-3-one

A mixture of8-bromo-9-(bromomethyl)-3,5,5-trimethyl-6,7-dihydro-5H-benzo[7]annulene(0.279 mmol, 1.0 equiv.), oxalic acid (1.67 mmol, 6 equiv.), PdCl2(0.013 mmol, 0.05 equiv.), dppe (0.013 mmol, 0.05 equiv.), TBACl (0.139mmol, 0.50 equiv.) in DMF and t-amyl alcohol in 1:1 ratio (0.5:0.5 ml)was placed in a pressurized reaction tube (5 ml) under conventionalheating (130° C.) for 20 hrs. After cooling to ambient temperature,water was added to the reaction mixture and extracted with ethyl acetate(3×10 ml). The combined organic layer was washed with water and driedover Na₂SO₄ and solvent was removed under reduced pressure. The viscousliquid obtained was purified by column chromatography on silica gel(60-120 mesh) using hexane:EtOAc (90:10) to afford the desired productas a white solid (97 mg, 40%).

¹H NMR (CDCl₃, 300 MHz, δ, ppm):1.35 (s, 6H), 2.15-2.11 (m, 2H), 2.37(s, 3H), 2.51-2.47 (t, 3H), 4.63 (s, 2H), 7.11-7.08 (d, J=6.99 Hz, 1H),7.23 (s, 1H), 7.33-7.30 (d, J=7.86 Hz, 1H); ¹³C NMR (CDCl₃, 75 MHz, δ,ppm): 20.1, 22.2, 27.5, 34.2, 36.6, 69.2, 125.1, 125.5, 125.7, 125.9,139.1, 150.2, 151.6, 173.6. ESI-MS: calcd for C₁₆H₁₈O₂[M+H]⁺ 243.1380,found 243.3711.

Example 3

General Procedure for the Synthesis of Formula II

Formula II (a) Synthesis of6,6,8-trimethyl-1,4,5,6-tetrahydrobenzo[3,4]cyclohepta[1,2-c]pyrrol-3(2H)-one

A mixture of8-bromo-9-(bromomethyl)-3,5,5-trimethyl-6,7-dihydro-5H-benzo[7]annulene(0.279 mmol, 1.0 equiv.), ammonium carbamate (1.1172 mmol, 4 equiv.),oxalic acid (1.67 mmol, 6 equiv.), PdCl₂ (0.013 mmol, 0.05 equiv.), dppe(0.013 mmol, 0.05 equiv.), TBACl (0.139 mmol, 0.50 equiv.) in DMF andt-amyl alcohol in 1:1 ratio (0.5:0.5 ml) was placed in a pressurizedreaction tube (5 ml) under conventional heating (130° C.) for 20 hrs.After cooling to ambient temperature, water was added to the reactionmixture and extracted with ethyl acetate (3×10 ml). The combined organiclayer was washed with water and dried over Na₂SO₄ and solvent wasremoved under reduced pressure. The viscous liquid obtained was purifiedby column chromatography on silica gel (60-120 mesh) using hexane:EtOAc(70:30) to afford the desired product as a yellow solid (20 mg, 30%). 1HNMR (CDCl3, 600 MHz, δ, ppm): 1.25 (s, 6H), 1.76-1.74 (t, J=6.66 Hz,2H), 2.31 (s, 3H), 2.50-2.46 (m, 2H), 4.25 (s, 2H), 7.07-7.06 (d, J=7.92Hz, 1H), 7.27 (s, 1H), 7.38-7.37 (d, J=7.98 Hz, 1H); 13C NMR (CDCl3, 150MHz, δ, ppm): 21.5, 24.1, 28.9, 36.4, 38.0, 47.9, 126.7, 127.2, 128.2,128.3, 133.4, 138.4, 146.6, 150.5, 174.1. ESI-MS: calcd for C₁₆H₁₉NO₂[M+H]⁺ 242.1539, found 242.1002.

Formula II (b) Synthesis of2-benzyl-6,6,8-trimethyl-1,4,5,6-tetrahydrobenzo[3,4]cyclohepta[1,2-c]pyrrol-3(2H)-one

A mixture of8-bromo-9-(bromomethyl)-3,5,5-trimethyl-6,7-dihydro-5H-benzo[7]annulene(0.279 mmol, 1.0 equiv.), phenylmethanamine (1.1172 mmol, 1.2 equiv.),oxalic acid (1.67 mmol, 6 equiv.), PdCl₂ (0.013 mmol, 0.05 equiv.), dppe(0.013 mmol, 0.05 equiv.), TBACl (0.139 mmol, 0.50 equiv.) in DMF andt-amyl alcohol in 1:1 ratio (0.5:0.5 ml) was placed in a pressurizedreaction tube (5 ml) under conventional heating (130° C.) for 20 hrs.After cooling to ambient temperature, water was added to the reactionmixture and extracted with ethyl acetate (3×10 ml). The combined organiclayer was washed with water and dried over Na₂SO₄ and solvent wasremoved under reduced pressure. The viscous liquid obtained was purifiedby column chromatography on silica gel (60-120 mesh) using hexane:EtOAc(70:30) to afford the desired product as a semisolid (25%). 1H NMR(CDCl3, 600 MHz) δ 1.34 (s, 6H), 1.90-1.88 (t, J=6.6 Hz, 2H), 2.36 (s,3H), 2.77-2.75 (t, J=6.5 Hz, 2H), 4.19 (s, 2H), 4.73 (s, 2H), 7.01-7.00(d, J=7.3 Hz, 1H), 7.18-7.16 (d, J=7.9 Hz, 1H), 7.32-7.28 (m, 4H),7.38-7.35 (m, 2H); 13C NMR (CDCl3, 150 MHz,) δ 22.01, 24.94, 29.38,36.68, 38.57, 46.68, 52.70, 127.23, 127.34, 127.91, 128.04, 128.21,128.56, 129.27, 134.08, 137.4, 137.91, 139.20, 144.77, 151.25, 172.68.ESI-MS: calcd. for C₂₃H₂₅NO [M+H]⁺=332.2009, found 331.8708.

Example 4

General Procedure for the Synthesis of Formula IV

Formula IV (a) Synthesis of3,5,5-trimethyl-5,6,7,9,10,11-hexahydro-12H-benzo[3,4]cycloheptal[1,2-b]naphthalene-12-one

A mixture of8-bromo-9-(bromomethyl)-3,5,5-trimethyl-6,7-dihydro-5H-benzo[7]annulene(0.279 mmol, 1.0 equiv.),3-amino-2-cyclohexen-1-one/3-aminocyclopent-2-en-1-one (0.33 mmol, 1.2equiv.), Pd(OAc)2 (0.055 mmol, 20 mol %), Xanthphos (0.055 mmol, 20 mol%), K₂CO₃ (0.55 mmol, 2 equvi.) in 2-methyl THF (1.5 ml) were placed inreaction tube (15 mL) at 90° C. for 12 h. After cooling the reactionmixture to ambient temperature, water was added and extracted with ethylacetate (10×3 times). The combined organic layer was washed with waterand dried over Na₂SO₄ and vacuum evaporated. The viscous liquid obtainedwhich was purified by column chromatography on silica gel (mesh 60-120)using the mixture of hexane: EtOAc (80:20) to obtained the desiredproduct as semisolid (38 mg, 45%).

¹H NMR (CDCl₃, 600 MHz, δ, ppm): 1.18 (s, 6H), 2.18-2.13 (m, 4H), 2.35(s, 3H), 2.66-2.62 (m, 2H), 2.76-2.71 (t, J=7.0 Hz, 2H), 3.12-3.08 (t,J=6.1 Hz, 2H), 7.11 (d, 1H), 7.21-7.16 (m, 2H), 8.10 (s, 1H); ¹³C NMR(CDCl₃, 150 MHz, δ, ppm): 22.07, 22.7, 29.6, 31.7, 35.5, 37.6, 38.5,47.3, 126.7, 127.1, 127.3, 130.6, 134.3, 137.3, 138.03, 145.8, 161.5,164.5, 198.2. ESI-MS: calcd for C₂₁H₂₃NO [M+H]⁺ 306.1852, found306.1964.

Formula IV (b) Synthesis of3,5,5,10-tetramethyl-5,6,7,9,10,11-hexahydro-12H-benzo[3,4]cyclohepta[1,2-b]quinolin-12-one

A mixture of8-bromo-9-(bromomethyl)-3,5,5-trimethyl-6,7-dihydro-5H-benzo[7]annulene(0.279 mmol, 1.0 equiv.),3-amino-2-cyclohexen-1-one/3-amino-6-methylcyclohex-2-en-1-one (0.33mmol, 1.2 equiv.), Pd(OAc)2 (0.055 mmol, 20 mol %), Xanthphos (0.055mmol, 20 mol %), K₂CO₃ (0.55 mmol, 2 equvi.) in 2-methyl THF (1.5 ml)were placed in reaction tube (15 mL) at 90° C. for 12 h. After coolingthe reaction mixture to ambient temperature, water was added andextracted with ethyl acetate (10×3 times). The combined organic layerwas washed with water and dried over Na₂SO₄ and vacuum evaporated. Theviscous liquid obtained which was purified by column chromatography onsilica gel (mesh 60-120) using the mixture of hexane: EtOAc (80:20) toobtained the desired product as semisolid (48%).

¹H NMR (CDCl₃, 600 MHz, δ, ppm): 1.23 (s, 6H), 1.24 (m, 3H), 2.27-2.24(m, 4H), 2.45 (s, 3H), 2.91-2.79 (m, 4H), 3.27-3.24 (m, 2H), 3.12-3.08(t, J=6.1 Hz, 2H), 7.20-7.19 (m, 1H), 7.28-7.27 (m, 1H), 8.18 (s, 1H);¹³C NMR (CDCl₃, 150 MHz, δ, ppm): 21.3, 21.6, 29.5, 31.7, 35.6, 37.6,40.6, 46.7, 47.3, 126.5, 126.7, 127.4, 130.6, 132.6, 134.3, 137.2,138.0, 145.8, 161.0, 164.6, 198.3. ESI-MS: calcd for C₂₂H₂₅NO [M+H]⁺320.2009, found 320.1964.

Example 5

General Procedure for the Synthesis of Formula VI

Formula VI (a) Synthesis of3,5,5-Trimethyl-9-((phenylsulfonyl)methyl)-6,7-dihydro-5H-benzo[7]annulene

A mixture of2,9,9-trimethyl-5-methylene-6,7,8,9-tetrahydro-5H-benzo[7]annulene (0.25mmol, 1.0 equiv.), sodium benzenesulfinate (0.3 mmol, 1.2 equiv.),K₂S₂O₈ (0.62 mmol, 2.5 equiv.), and I₂ (0.3 mmol, 1.2 equiv.), inACN:H₂O (1:1) were placed in reaction tube (5 mL) at room temperaturefor 12 h. After completion of reaction, a saturated solution of sodiumthiosulfate was added to the reaction mixture and extracted with ethylacetate (10×3 times). The combined organic layer was washed with waterand dried over Na₂SO₄ and vacuum evaporated. The viscous liquid obtainedwas purified by column chromatography on silica gel (mesh 60-120) usingthe mixture of hexane: EtOAc (80:20) to obtained the desired product assemisolid (71 mg, 84%).

¹H NMR (CDCl₃, 600 MHz) δ 1.34 (s, 6H), 1.96-1.99 (t, J=7.2 Hz, 2H),2.10-2.12 (t, J=6.9 Hz, 2H), 2.34 (s, 3H), 4.22 (s, 2H), 6.24-6.26 (t,J=6.4 Hz, 1H), 6.95 (d, J=7.8 Hz, 1H), 7.20 (d, J=8.1 Hz, 2H), 7.51-7.53(t, J=7.8 Hz, 2H), 7.61-7.64 (m, 1H), 7.90 (d, J=7.5 Hz, 2H); ¹³C NMR(CDCl₃, 150 MHz) δ 21.3, 27.0, 30.8, 38.0, 46.5, 63.7, 126.4, 126.5,128.1, 128.3, 128.4, 129.0, 133.4, 134.4, 136.6, 137.0, 139.7, 148.0.ESI-MS: calcd. for C₂₁H₂₄O₂S [M+H]⁺ 341.1575, found 341.1565.

Formula VI(b) Synthesis of3,5,5-trimethyl-9-(tosylmethyl)-6,7-dihydro-5H-benzo[7]annulene

A mixture of2,9,9-trimethyl-5-methylene-6,7,8,9-tetrahydro-5H-benzo[7]annulene (0.25mmol, 1.0 equiv.), sodium 4-methylbenzenesulfinate (0.3 mmol, 1.2equiv.), K₂S₂O₈ (0.62 mmol, 2.5 equiv.), and I₂ (0.3 mmol, 1.2 equiv.),in ACN:H₂O (1:1) were placed in reaction tube (5 mL) at room temperaturefor 12 h. After completion of reaction, a saturated solution of sodiumthiosulfate was added to the reaction mixture and extracted with ethylacetate (10×3 times). The combined organic layer was washed with waterand dried over Na₂SO₄ and vacuum evaporated. The viscous liquid obtainedwas purified by column chromatography on silica gel (mesh 60-120) usingthe mixture of hexane: EtOAc (80:20) to obtained the desired product assemisolid (64 mg, 72%).

1H NMR (CDCl₃, 600 MHz) δ 1.35 (s, 6H), 1.97-2.00 (t, J=7.2 Hz, 2H),2.11-2.14 (m, 2H), 2.35 (s, 3H), 2.45 (s, 3H), 4.21 (s, 2H), 6.24-6.27(t, J=6.5 Hz, 1H), 6.96-6.97 (d, J=7.3 Hz, 1H), 7.21-7.24 (t, J=8.5 Hz,2H), 7.31-7.32 (d, J=8.1 Hz, 2H), 7.78-7.79 (d, J=8.2 Hz, 2H); 13C NMR(CDCl₃, 150 MHz) δ 21.49, 21.65, 27.12, 30.92, 38.09, 46.69, 63.88,126.46, 126.62, 128.38, 128.43, 128.60, 129.71, 134.63, 136.57, 136.79,136.85, 144.50, 148.08. ESI-MS: calcd. For C₂₂H₂₆O₂S [M+H]⁺ 355.1732,found 355.1720.

Formula VI (c) Synthesis of3,5,5-Trimethyl-9-((methylsulfonyl)methyl)-6,7-dihydro-5H-benzo[7]annulene

A mixture of2,9,9-trimethyl-5-methylene-6,7,8,9-tetrahydro-5H-benzo[7]annulene (0.25mmol, 1.0 equiv.), sodium methanesulfinate (0.3 mmol, 1.2 equiv.),K₂S₂O₈ (0.62 mmol, 2.5 equiv.), and 12 (0.3 mmol, 1.2 equiv.), inACN:H₂O (1:1) were placed in reaction tube (5 mL) at room temperaturefor 12 h. After completion of reaction, a saturated solution of sodiumthiosulfate was added to the reaction mixture and extracted with ethylacetate (10×3 times). The combined organic layer was washed with waterand dried over Na₂SO₄ and vacuum evaporated. The viscous liquid obtainedwas purified by column chromatography on silica gel (mesh 60-120) usingthe mixture of hexane: EtOAc (60:40) to obtained the desired product assemisolid (42 mg, 60%).

¹H NMR (CDCl₃, 600 MHz) δ 1.40 (s, 6H), 2.00-2.02 (t, J=7.0 Hz, 2H),2.20-2.23 (m, 2H), 2.37 (s, 3H), 2.71 (s, 3H), 4.19 (s, 2H), 6.40-6.42(t, J=6.2 Hz, 1H), 7.09 (d, J=7.8 Hz, 1H), 7.27 (s, 1H), 7.32 (d, J=7.8Hz, 1H); ¹³C NMR (CDCl₃, 150 MHz) δ 21.39, 27.18, 30.58, 38.09, 41.10,45.98, 62.80, 126.74, 127.04, 127.96, 128.12, 133.68, 137.12, 137.40,148.54. ESI-MS: calcd. for C₁₆H₂₂O₂S [M+H]⁺ 279.1419, found 279.1410.

Example 6 General Procedure for the Synthesis of Formula V

Synthesis of3,5,5,10-tetramethyl-6,7-dihydro-5H-benzo[3,4]cyclohepta[1,2-b]quinolin-12-ol

A mixture of3,5,5,10-tetramethyl-5,6,7,9,10,11-hexahydro-12H-benzo[3,4]cyclohepta[1,2-b]quinolin-12-one(1.0 equiv.), oxidant (0.3 mmol, 2 equiv.) in solvent (1:1) were placedin reaction tube (5 mL) for 12 h. After cooling the reaction mixture toambient temperature, water was added and extracted with ethyl acetate(10×3 times). The combined organic layer was washed with water and driedover Na₂SO₄ and vacuum evaporated. The viscous liquid obtained which waspurified by column chromatography on silica gel (mesh 60-120) using themixture of hexane: EtOAc (70:30) to obtained the desired product.

Experimental Procedures

-   -   I. Determination of affinity and kinetic constants for PI3K        (120y)    -   II. Determination of affinity of PI3K (120y) inhibitors        (KD/IC50) through inhibition in solution assay of kinase PI3K        (120y) by various compounds of general formula I-VI    -   III. Determination of affinity and kinetic constants for        MAPKAPK2    -   IV. Determination of affinity of MK2 inhibitors (IC50) through        inhibition in solution assay of kinase MK2 by various compounds        of general formula I-VI

Determination of Affinity and Kinetic Constants for PI3K (120y)

Immobilization buffer scouting was performed for PI3K (120y) to find asuitable buffer for immobilization. It was found that 10 mM sodiumacetate pH 5.5 buffer is best suitable for the immobilization.Immobilization of the PI3K (120y) kinase was performed at 25° C.temperature using concentration of Kinase 5 μg/ml with a flow rate of 10μl/min and 1000 sec of contact time. Successful immobilization of 9191.2RU of kinase PI3K (120y) over flow cell 4 of sensor surface CM5 by aminecoupling was achieved.

After immobilization, kinetics screening for binding of compounds withPI3K (120y) was performed with the protocol recommended as per BiacoreAssay Handbook (protocol: kinetic screening using a single concentrationof compounds, concentration used: 50 μMin 1×PBS with 5% DMSO, runningbuffer: 1×PBS with 5% DMSO, temperature: 25° C., buffer blanks: 1×PBSwith 5% DMSO wizard used: Kinetics/Affinity). Compound K were preparedin 50 μM concentration in 1× PBS with 5% DMSO and placed in samplecompartment at 25° C. These were passed over the sensor through flowcell 1 (reference-blank) and 2 (active-immobilized with kinase) in flowrate of 30 μl/min at 25° C. Binding responses were seen in real time forFc 2 (active), Fc1 (reference) and Fc 2-1 (reference subtracted).Solvent correction was done for DMSO (5% in PBS) Data was analyzed usingBiacore T200 evaluation software v 3.1 and fitting was done using 1:1binding model. Also, detailed kinetics characterization was performedfor compounds of general formula I.

Compounds of general formula II showed promising binding and kineticsprofiles as shown in FIG. 1 , FIG. 2 , and Table 1 FIG. 1 depicts theon-rate and off-rate of binding of evaluated compounds of generalformula II to PI3K (120y). Plotting association rate constant ka againstdissociation rate constant kd (here on logarithmic scales) creates aplot where the affinity is represented by diagonal lines. Compounds onthe same diagonal have the same affinity but differ in kinetics. FIG. 2is the Graphical representation of the on-rate and off-rate of bindingof evaluated compounds of general formula II to PI3K (120y).

Table 1 represents the numerical values of binding (on-rate, off-rateand affinity of binding) of evaluated compounds of general formula I toPI3K (120y). KD represents the equilibrium constant of binding in Molarvalue.

TABLE 1 Sample Ka (1/Ms) Kd (1/s) KD (M) Sample 5 9.75E+03 2.38E−032.44E−07 Sample 7 1.70E+04 2.46E−01 1.45E−05 Sample 8 9.88E+03 2.41E−012.44E−05 Sample 9 1.08E+04 2.31E−01 2.14E−05 Sample 11 1.44E+04 1.51E−011.05E−05 Sample 12 4.46E+04 9.48E−03 2.13E−07 Sample 16 3.16E+045.54E−02 1.75E−06

Detailed kinetics characterization of selected compounds (Compoundshaving general formula II that is pyrolone fused benzocycloheptene) forbinding to PI3K (120y) were performed using the protocol recommended asper Biacore Assay Handbook (protocol: kinetic analysis using singlecycle kinetics method, concentrations used: 10, 5, 2.5, 1.25, 0.625 μMin 1×PBS with 5% DMSO, running buffer: 1×PBS with 5% DMSO, temperature:25° C. Buffer blanks: 1×PBS with 5% DMSO, wizard used:Kinetics/Affinity). Compound (Compounds having general formula II thatis pyrolone fused benzocycloheptene) were prepared in 10 μMconcentration in 1× PBS with 5% DMSO and serial diluted to make 5, 2.5,1.25 and 0.625 μM concentration. They were placed in sample compartmentat 25° C. These were passed over the sensor through flow cell 1(reference-blank) and 2 (active-immobilized with kinase) in flow rate of30 μl/min at 25° C. using single cycle kinetics injection method.Binding responses were seen in real time for Fc2 (active), Fc1(reference) and Fc 2-1 (reference subtracted). Solvent correction wasdone for DMSO (5% in PBS). Data was analyzed using Biacore T200evaluation software v 3.1. Fitting of data is done using 1:1 bindingmodel and kinetics constants as well as equilibrium kinetics constant ofdissociation (KD) was also determined.

Compounds of general formula II showed effective inhibitory potentialagainst PI3K (120y) with IC50 ranging in between 2-2.5 μM as provided inFIG. 3 , and Table 2. FIG. 3 represents the dose dependent inhibition ofthe PI3K (120y) by compounds of general formula II (5, 7, 8, 9, 12 and16). Table 2: Table represents the extrapolated IC50 values of thecompounds of general formula I against the PI3K(120y) from the graphs.

TABLE 2 S. No. Ligand (Small Molecules) IC50 1 Ligand 5  2057 nM 2Ligand 7  1958 nM 3 Ligand 8  2173 nM 4 Ligand 9  1999 nM 6 Ligand 122342 nM 7 Ligand 16 2093 nM

Example 7 Determination of Affinity and Kinetic Constants for Mk2

Immobilization buffer scouting was performed for MK2 to find suitablebuffer for immobilization and it was found that 10 mM Sodium acetate pH5.5 buffer is best suitable for the immobilization. Immobilization ofthe MK2 kinase was performed at 25° C. temperature using concentrationof Kinase 10 μg/ml with a flow rate of 10 μl/min and 1000 sec of contacttime. Successful immobilization of 11119.6 RU of kinase MK2 over flowcell 4 of sensor surface CM5 by amine coupling was achieved.

After immobilization, kinetics screening for binding of compounds withMK2 was performed with the protocol recommended as per Biacore AssayHandbook (protocol: kinetic screening using a single concentration ofcompounds, concentration used: 50 μM in 1×PBS with 5% DMSO, runningbuffer: 1×PBS with 5% DMSO, temperature: 25° C., buffer blanks: 1×PBSwith 5% DMSO wizard used: Kinetics/Affinity). Compound (Compounds havinggeneral formula II that is pyrolone fused benzocycloheptene) wereprepared in 50 μM concentration in 1× PBS with 5% DMSO and placed in asample compartment at 25° C. These were passed over the sensor throughflow cell 3 (reference-blank) and 4 (active-immobilized with kinase) inflow rate of 30 μl/min at 25° C. Binding responses were seen in realtime for Fc 4 (active), Fc3 (reference) and Fc 4-3 (referencesubtracted). Solvent correction was done for DMSO (5% in PBS). Data wasanalyzed using Biacore T200 evaluation software v 3.1 and fitting wasdone using 1:1 binding model.

Compounds of general formula II showed promising binding and kineticsprofiles. Out of them, on the basis of their binding and kineticprofiles, 9 most promising lead compounds of general formula II alongwith one marketed MK2-inhibitor compound (PF 3644022) were selected forfurther in-solution inhibition assay. The results are provided in FIG. 4, FIG. 5 , and Table 3. FIG. 4 depicts the on-rate and off-rate ofbinding of evaluated compounds of general formula II to MK2. Plottingassociation rate constant ka against dissociation rate constant kd (hereon logarithmic scales) creates a plot where the affinity is representedby diagonal lines. Compounds on the same diagonal have the same affinitybut differ in kinetics. FIG. 5 is the graphical representation of theon-rate and off-rate of binding of evaluated compounds of generalformula II to MK2. Table 3 represents the numerical values of binding(on-rate, off-rate and affinity of binding) of evaluated compounds ofgeneral formula II to MK2. KD represents the equilibrium constant ofbinding in Molar value.

TABLE 3 Sample Ka (1/Ms) Kd (1/s) KD (M) Sample 1 2.11E+03 1.28E−016.08E−05 Sample 2 3.75E+03 9.30E−02 2.48E−05 Sample 3 2.55E+03 3.63E−021.42E−05 Sample 4 4.12E+03 6.61E−02 1.61E−05 Sample 5 1.11E+00 2.82E−032.53E−03 Sample 6 1.77E+01 5.87E−02 3.32E−03 Sample 7 3.46E+03 1.36E−013.94E−05 Sample 8 3.42E+03 1.24E−01 3.63E−05 Sample 9 3.57E+03 6.73E−021.89E−05 Sample 11 2.41E+03 1.54E−01 6.40E−05 Sample 12 2.65E+035.39E−02 2.04E−05 Sample 13 3.32E+03 7.88E−02 2.37E−05 Sample 143.26E+03 8.61E−02 2.64E−05 Sample 15 1.94E+03 1.09E−01 5.64E−05 Sample16 1.70E+03 1.67E−01 9.81E−05

Detailed kinetics characterization of selected compounds binding to MK2were performed using the protocol recommended as per Biacore AssayHandbook (protocol: kinetic analysis using single cycle kinetics method,concentrations used: 30, 15, 7.5, 3.75, 1.87 μMin 1×PBS with 5% DMSO,running buffer: 1×PBS with 5% DMSO, temperature: 25° C., buffer blanks:1×PBS with 5% DMSO, wizard used: Kinetics/Affinity). Compounds wereprepared in 30 μM concentration in 1× PBS with 5% DMSO and serialdiluted to make 15, 7.5, 3.75 and 1.87 μM concentration. They wereplaced in a sample compartment at 25° C. These were passed over thesensor through flow cell 3 (reference-blank) and 4 (active-immobilizedwith kinase) in flow rate of 30 μl/min at 25° C. using single cyclekinetics injection method. Binding responses were observed in real timefor Fc 4 (active), Fc3 (reference) and Fc 4-3 (reference subtracted).Solvent correction was done for DMSO (5% in PBS). Data was analyzedusing Biacore T200 evaluation software v 3.1 and fitting of data wasdone using 1:1 binding model and kinetics constants as well asequilibrium kinetics constant of dissociation (KD) was also determined.

Compounds of general formula II showed effective inhibitory potentialagainst MK2 with IC50 between ranging in between 526-709 nM. The valuesare in well competence with IC50 value of marketed MK2-inhibitor (585nM) as provided in FIG. 6 , and Table 4.

FIG. 6 : Graphs represent the dose dependent inhibition of the MK2 bycompounds of general formula II (1, 3, 6, 8, 12, 13, 14, 15 and 17 alongwith marketed MK2-inhibitor PF 3644022. Table 4 represents theextrapolated IC50 values of the compounds of general formula II againstMK2 from the graphs.

TABLE 4 S. No. Ligand (Small Molecules) IC50 1 Ligand 1  650 nM 2 Ligand3  526 nM 3 Ligand 6  550 nM 4 Ligand 8  525 nM 5 Ligand 12 709 nM 6Ligand 13 578 nM 7 Ligand 14 545 nM 8 Ligand 15 531 nM 9 Ligand 17 596nM 10 Marketed MK2 Inhibitor (PF 3644022) 585 nM

Advantages of the Invention

-   -   The main scaffold of benzocycloheptene comes from natural        precursor. Therefore, use of natural precursor reduces the        number of steps for the synthesis of the compounds.    -   The cost of obtaining plant derived precursor is very low, as        the plant is abundant in nature, hence the overall process is        economic.    -   The process is also sustainable as 60-80% of the molecules come        from natural precursor and only 40-20% molecule were used        through synthetic process.    -   The compounds of general formula I-VI are less toxic.    -   The compound of general formula I-VI are applicable for        treatment of PI3K and MK2 related disorder.

11. A compound of formula (I), (II), (III), (IV), (V), or (VI):

or a pharmaceutical acceptable salt or enantiomer thereof, wherein, ineach of formulas (I), (II), (III), (IV), (V), and (VI): Y is selectedfrom the group consisting of carbon, nitrogen, oxygen, and sulfur; Z isselected from the group consisting of —H, C1-C6 alkyl, C1-C6 aryl,halogen, oxygen, nitrogen, and sulfur; R¹ is selected from the groupconsisting of —H, —OH, N-substituted benzylamine, N-substituted aniline,S-substituted thiophenes, carboxylic acids, and halogen; R² is selectedfrom the group consisting of —H, C1-C6 alkyl, C1-C6 aryl, halogen, C1-C6amine, and C1-C6 carbonyl groups; R³ is selected from the groupconsisting of —H, C1-C6 alkyl, C1-C6 aryl, and thiophenes; X is selectedfrom the group consisting of carbon, nitrogen, oxygen, and sulfur; R⁴ isselected from the group consisting of —H, C1-C6 alkyl, C1-C6 aryl,halogen, and aldehyde-substituted benzenesulfonamides; R⁵ is selectedfrom the group consisting of —H, C1-C6 alkyl, C1-C6 aryl, halogen, andC1-C6 carbonyl group; and R⁶ is selected from the group consisting of—H, C1-C6 alkyl, C1-C6 aryl, and C1-C6 amine group.
 12. The compound ofclaim 11, wherein the compound has formula (I), where: R¹ is selectedfrom the group consisting of —H, —OH, N-substituted benzylamine,N-substituted anilines, carboxylic acids, and halogens, R² is selectedfrom the group consisting of —H, C1-C6 alkyl, C1-C6 aryl, halogen, C1-C6amine, and C1-C6 carbonyl group; and R³, R⁴, and R⁵ are eachindependently selected from the group consisting of —H, C1-C6 alkyl,C1-C6 aryl, halogen, and C1-C6 carbonyl group.
 13. The compound of claim11, wherein the compound has formula (II), where: R¹ is selected fromthe group consisting of —H, OH, N-substituted benzylamine, N-substitutedaniline, carboxylic acids, and halogen, R² is selected from the groupconsisting of —H, C1-C6 alkyl, C1-C6 aryl, halogen, C1-C6 amine, andC1-C6 carbonyl; and Y is selected from the group consisting of carbon,oxygen, nitrogen, and sulfur.
 14. The compound of claim 11, wherein thecompound has formula (III), where: R¹ is selected from the groupconsisting of —H, —OH, benzylamine, aniline, carboxylic acids, andhalogen; R² is selected from the group consisting of —H, C1-C6 alkyl,C1-C6 aryl, halide, C1-C6 amine, and C1-C6 carbonyl; and Z is selectedfrom the group consisting of —H, C1-C6 alkyl, C1-C6 aryl, halogen,oxygen, nitrogen, and sulfur.
 15. The compound of claim 11, wherein thecompound has formula (IV), where: R¹ is selected from the groupconsisting of —H, —OH, N-substituted benzylamine, N-substituted aniline,carboxylic acids, and halogen, R² is selected from the group consistingof —H, C1-C6 alkyl, C1-C6 aryl, halogen, C1-C6 amine, and C1-C6carbonyl; and n is 5 or
 6. 16. The compound of claim 11, wherein thecompound has the formula (V), where: R¹ is selected from the groupconsisting of —H, —OH, N-substituted benzylamine, N-substituted aniline,carboxylic acids, and halogen; R² is selected from the group consistingof —H, C1-C6 alkyl, C1-C6 aryl, halogen, C1-C6 amine, and C1-C6 carbonylgroup; n is from 5 to 6; and R³ is selected from the group consisting of—H, C1-C6 alkyl, C1-C6 aryl, and halogen.
 17. The compound of claim 11,wherein the compound has formula (VI), where: R¹ is selected from thegroup consisting of —H, —OH, N-substituted benzylamine, N-substitutedaniline, carboxylic acids, and halogen; R² is selected from the groupconsisting of —H, C1-C6 alkyl, C1-C6 aryl, halogen, C1-C6 amine, andC1-C6 carbonyl; R⁵ is selected from the group consisting of hydrogen,C1-C6 alkyl, halogen, sulfur, C1-C6 amine, and C1-C6 aryl; and R⁶ isselected from the group consisting of —H, C1-C6 alkyl, C1-C6 aryl, andC1-C6 amine.
 18. A pharmaceutical formulation comprising the compoundaccording to claim 11 and a pharmaceutically acceptable adjuvant,diluent, or carrier.
 19. A method for treating type-2 diabetes, fever,inflammation, cancer, ulcers, or for stimulating or depressing thecentral nervous system in a subject, the method comprising administeringthe compound according to claim 11 to the subject.
 20. A method ofinhibiting PI3K and MK2 mediated activity in a subject, the methodcomprising administering the compound according to claim 11 to thesubject.